The long range goal of the proposed research is to understand the functional relationships of the secretory and systemic immune systems in fish. Two general hypotheses are to be tested: 1) fish have distinct antibody (Ab) isotypes which have different biological properties and different immune functions and 2) fish have a secretory immune system which can function independently of the systemic system. The objectives proposed are four-fold. Basic studies detailed in this proposal have shown that fish have different classes, subclasses, and subgroups of immunoglobulin (Ig) heavy (H) and light (L) chains. Fish have been found to have three antigenically and structurally distinct H chain subclasses. These H chain classes represent about 60% of the total Igs in serum. The first objective is to define the remaining subclasses with monoclonal Abs and then to compare the structure of the newly defined subclasses with those now identified by comparative peptide mapping analyses. Fish have two classes of Ig L chains, reminiscent of k and Lambda in higher vertebrates. The second objective is to obtain primary sequence information of defined regions of interest to establish the relatedness of these proteins to the mammalian forms and to gain structural insight into phylogenetic divergence of Ig proteins. The third objective is to determine if the different subclasses have different biological properties important in immune defense. Antibodies are known to function in cooperation with the complement system and phagocytic cells to provide basic immunity. These studies will address the ability of the different Ig subclasses to bind complement by the direct pathway, to bind to macrophage surfaces and thereby promote phagocytosis, to be passively transferred to the egg and thereby provide immune function to the young embryo, and to determine the metabolic relationships of the Igs found in the systemic and secretory systems. The last objective is to determine the immune Ab profile of fish immunized with defined haptens and carriers. Fish will be immunized by parenteral and nonparenteral routes and the level and isotype of the elicited Ab defined in the serum, cutaneous mucus, and in the gut. These studies will provide important basic information on the functional relationship of the secretory and systemic immune system in fish, provide a phylogenetic basis for Ig isotype divergence, and provide economically important information to a rapidly growing aquaculture industry.